The Plasmodium falciparum–CD36 Interaction Is Modified by a Single Amino Acid Substitution in CD36

Blood ◽  
1998 ◽  
Vol 92 (5) ◽  
pp. 1814-1819 ◽  
Author(s):  
Lena Serghides ◽  
Ian Crandall ◽  
Eric Hull ◽  
Kevin C. Kain
Keyword(s):  
Plasmodium Falciparum ◽  
Molecular Mechanisms ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid ◽  
Directed Mutagenesis ◽  
Binding Characteristics ◽  
Ofplasmodium Falciparum ◽  
Ph Dependent ◽  
American Society ◽  
Ph Range

CD36 is an 88-kD glycoprotein involved in the cytoadherence ofPlasmodium falciparum–parasitized erythrocytes (PE) to endothelial cells. The molecular mechanisms involved in CD36-dependent cytoadherence were examined by expressing three CD36 homologues (human, murine, and rat) in COS-7 cells and observing their PE-binding characteristics over a pH range of 6.0 to 7.4 and following iodination of these receptors. PE binding to human CD36 was pH dependent, with peak binding at pH 6.8 to 7.0, and binding was unaffected by iodination. In contrast, PE adherence to murine and rat CD36 was insensitive to changes in pH, and iodination significantly reduced binding. We further show that the differences observed in the binding phenotype of human and rodent CD36 can be attributed to a single residue. Site-directed mutagenesis of the histidine at position 242 of human CD36 to tyrosine (found in rodent CD36) conferred the binding phenotype of rodent CD36 onto human CD36. Furthermore, substitution of the tyrosine at position 242 of rat CD36 for histidine conferred the binding phenotype of human CD36 onto rat CD36. These findings suggest that residue 242 is part of, or important to the conformation of, the PE-binding domain of CD36. © 1998 by The American Society of Hematology.

The Plasmodium falciparum–CD36 Interaction Is Modified by a Single Amino Acid Substitution in CD36

Blood ◽  
1998 ◽  
Vol 92 (5) ◽  
pp. 1814-1819 ◽  
Author(s):  
Lena Serghides ◽  
Ian Crandall ◽  
Eric Hull ◽  
Kevin C. Kain
Keyword(s):  
Plasmodium Falciparum ◽  
Molecular Mechanisms ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid ◽  
Directed Mutagenesis ◽  
Binding Characteristics ◽  
Ofplasmodium Falciparum ◽  
Ph Dependent ◽  
American Society ◽  
Ph Range

Abstract CD36 is an 88-kD glycoprotein involved in the cytoadherence ofPlasmodium falciparum–parasitized erythrocytes (PE) to endothelial cells. The molecular mechanisms involved in CD36-dependent cytoadherence were examined by expressing three CD36 homologues (human, murine, and rat) in COS-7 cells and observing their PE-binding characteristics over a pH range of 6.0 to 7.4 and following iodination of these receptors. PE binding to human CD36 was pH dependent, with peak binding at pH 6.8 to 7.0, and binding was unaffected by iodination. In contrast, PE adherence to murine and rat CD36 was insensitive to changes in pH, and iodination significantly reduced binding. We further show that the differences observed in the binding phenotype of human and rodent CD36 can be attributed to a single residue. Site-directed mutagenesis of the histidine at position 242 of human CD36 to tyrosine (found in rodent CD36) conferred the binding phenotype of rodent CD36 onto human CD36. Furthermore, substitution of the tyrosine at position 242 of rat CD36 for histidine conferred the binding phenotype of human CD36 onto rat CD36. These findings suggest that residue 242 is part of, or important to the conformation of, the PE-binding domain of CD36. © 1998 by The American Society of Hematology.

scholarly journals Polymerase and hydrolase activities of Bacillus subtilis levansucrase can be separately modulated by site-directed mutagenesis

1991 ◽  
Vol 279 (1) ◽  
pp. 35-41 ◽  
Author(s):  
R Chambert ◽  
M F Petit-Glatron
Keyword(s):  
Bacillus Subtilis ◽  
Intermediate Formation ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid ◽  
Chain Elongation ◽  
Directed Mutagenesis ◽  
Water Mixture ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Sucrose Hydrolysis

The levansucrase (sucrose:2,6-beta-D-fructan 6-beta-D-fructosyltransferase, EC 2.4.1.10) structural gene from a Bacillus subtilis mutant strain displaying a low polymerase activity was sequenced. Only one missense mutation changing Arg331 to His was responsible for this modified catalytic property. From this allele we created new mutations by directed mutagenesis, which modified the charge and polarity of site 331. Examination of the kinetics of the purified levansucrase variants revealed that transfructosylation activities are affected differently by the substitution chosen. His331→Arg completely restored the properties of the wild-type enzyme. The most striking feature of the other variants, namely Lys331, Ser331 and Leu331, was that they lost the ability of the wild-type enzyme to synthesize levan from sucrose alone. They were only capable of catalysing the first step of levan chain elongation, which is the formation of the trisaccharide ketose. The variant His331→Lys presented a higher kcat. for sucrose hydrolysis than the wild-type, and only this hydrolase activity was preserved in a solvent/water mixture in which the wild-type acted as a true polymerase. The two other substitutions reduced the efficiency of transfructosylation activities of the enzyme via the decrease of the rate of fructosyl-enzyme intermediate formation. For all variants, the sucrose affinity was slightly affected. This strong modulation of the enzyme specificities from a single amino acid substitution led us to postulate the hypothesis that bacterial levansucrases and plant fructosyltransferases involved in fructan synthesis may possess a common ancestral form.

scholarly journals Mammalian inositol monophosphatase: the identification of residues important for the binding of Mg2+ and Li+ ions using fluorescence spectroscopy and site-directed mutagenesis

1993 ◽  
Vol 296 (3) ◽  
pp. 811-815 ◽  
Author(s):  
M G Gore ◽  
P Greasley ◽  
G McAllister ◽  
C I Ragan
Keyword(s):  
Fluorescence Spectroscopy ◽  
Specific Activity ◽  
Fluorescence Emission ◽  
Site Directed Mutagenesis ◽  
Native Enzyme ◽  
Mutant Enzyme ◽  
Directed Mutagenesis ◽  
Inositol Monophosphatase ◽  
Fluorescence Emission Intensity ◽  
Ph Dependent

The fluorescence properties of residue Trp-219 in inositol monophosphatase are sensitive to the ionization of neighbouring groups. The pH-dependent changes in the fluorescence emission intensity and wavelength of maximum emission appear to arise as the result of two separate ionizations in the proximity of Trp-219, namely due to the ionization of His-217 and Cys-218. By studying the curve of fluorescence intensity against pH, given by the mutants Cys-218→Ala or His-217→Gln, the pK of His-217 was determined to be 7.54 and the pK of Cys-218 was estimated to be about 8.2. These mutants have altered kinetic parameters for catalytic Mg2+ ions and inhibitory Mg2+ and Li+ ions. The Cys-218→Ala mutant enzyme is not subject to inhibition by concentrations of Mg2+ ions up to 400 mM and has a specific activity of 156% of the maximum obtainable activity of the native enzyme. The His-217→Gln mutant enzyme shows reduced sensitivity to inhibition by Mg2+ and Li+ ions, and has a specific activity of 110% of that obtainable for the native enzyme.

scholarly journals Isolation from Ochrobactrum anthropi of a Novel Class II 5-Enopyruvylshikimate-3-Phosphate Synthase with High Tolerance to Glyphosate

2010 ◽  
Vol 76 (17) ◽  
pp. 6001-6005 ◽  
Author(s):  
Yong-Sheng Tian ◽  
Ai-Sheng Xiong ◽  
Jing Xu ◽  
Wei Zhao ◽  
Feng Gao ◽  
...  
Keyword(s):  
Genomic Library ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid ◽  
Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Construction Process ◽  
Ochrobactrum Anthropi ◽  
Gene Encoding ◽  
Phosphate Synthase

ABSTRACT Applying the genomic library construction process and colony screening, a novel aro A gene encoding 5-enopyruvylshikimate-3-phosphate synthase from Ochrobactrum anthropi was identified, cloned, and overexpressed, and the enzyme was purified to homogeneity. Furthermore, site-directed mutagenesis was employed to assess the role of single amino acid residues in glyphosate resistance.

scholarly journals Schistosome-derived omega-1 drives Th2 polarization by suppressing protein synthesis following internalization by the mannose receptor

2012 ◽  
Vol 209 (10) ◽  
pp. 1753-1767 ◽  
Author(s):  
Bart Everts ◽  
Leonie Hussaarts ◽  
Nicole N. Driessen ◽  
Moniek H.J. Meevissen ◽  
Gabriele Schramm ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Protein Synthesis ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Mannose Receptor ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Th2 Polarization ◽  
Th2 Responses ◽  
Egg Antigen

Omega-1, a glycosylated T2 ribonuclease (RNase) secreted by Schistosoma mansoni eggs and abundantly present in soluble egg antigen, has recently been shown to condition dendritic cells (DCs) to prime Th2 responses. However, the molecular mechanisms underlying this effect remain unknown. We show in this study by site-directed mutagenesis of omega-1 that both the glycosylation and the RNase activity are essential to condition DCs for Th2 polarization. Mechanistically, we demonstrate that omega-1 is bound and internalized via its glycans by the mannose receptor (MR) and subsequently impairs protein synthesis by degrading both ribosomal and messenger RNA. These experiments reveal an unrecognized pathway involving MR and interference with protein synthesis that conditions DCs for Th2 priming.

scholarly journals A Single Amino Acid Substitution in a Mannosyltransferase, WbdA, Converts the Escherichia coli O9 Polysaccharide into O9a: Generation of a New O-Serotype Group

2000 ◽  
Vol 182 (9) ◽  
pp. 2567-2573 ◽  
Author(s):  
Nobuo Kido ◽  
Hidemitsu Kobayashi
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Substitution ◽  
Site Directed Mutagenesis ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Directed Mutagenesis ◽  
E Coli ◽  
Chimeric Genes ◽  
Functional Studies

ABSTRACT wbdA is a mannosyltransferase gene that is involved in synthesis of the Escherichia coli O9a polysaccharide, a mannose homopolymer with a repeating unit of 2-αMan-1,2-αMan-1,3-αMan-1,3-αMan-1. The equivalent structural O polysaccharide in the E. coli O9 andKlebsiella O3 strains is 2-αMan-1,2-αMan-1,2-αMan-1,3-αMan-1,3-αMan-1, with an excess of one mannose in the 1,2 linkage. We have cloned wbdAgenes from these O9 and O3 strains and shown by genetic and functional studies that wbdA is the only gene determining the O-polysaccharide structure of O9 or O9a. Based on functional analysis of chimeric genes and site-directed mutagenesis, we showed that a single amino acid substitution, C55R, in WbdA of E. coli O9 converts the O9 polysaccharide into O9a. DNA sequencing revealed the substitution to be conserved in other E. coli O9a strains. The reverse substitution, R55C, in WbdA of E. coli O9a resulted in lipopolysaccharide synthesis showing no ladder profile instead of the conversion of O9a to O9. This suggests that more than one amino acid substitution in WbdA is required for conversion from O9a to O9.

Site-directed mutagenesis provides insights into the selective binding of trityl derivatives to Plasmodium falciparum dUTPase

2011 ◽  
Vol 46 (8) ◽  
pp. 3309-3314 ◽  
Author(s):  
Eliseo Recio ◽  
Alexander Musso-Buendía ◽  
Antonio E. Vidal ◽  
Gian Filippo Ruda ◽  
Ganasan Kasinathan ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Selective Binding
Sign in / Sign up

Export Citation Format

Share Document